Journal article
A Sensitive Alternative for MicroRNA In Situ Hybridizations Using Probes of 2'-O-Methyl RNA + LNA
The use of short, high-affinity probes consisting of a combination of DNA and locked nucleic acid (LNA) has enabled the specific detection of microRNAs (miRNAs) by in situ hybridization (ISH). However, detection of low–copy number miRNAs is still not always possible. Here the authors show that probes consisting of 2'-O-methyl RNAs (2OMe) and LNA at every third base (2:1 ratio), under optimized hybridization conditions, excluding yeast RNA from the hybridization buffer, can provide superior performance in detection of miRNA targets in terms of sensitivity and signal-to-noise ratio compared to DNA + LNA probes.
Furthermore, they show that hybridizations can be performed in buffers of 4M urea instead of 50% formamide, thereby yielding an equally specific but nontoxic assay. The use of 2OMe + LNA–based probes and the optimized ISH assay enable simple and fast detection of low–copy number miRNA targets, such as miR-130a in mouse brain.
Language: | English |
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Publisher: | SAGE Publications |
Year: | 2011 |
Pages: | 661-672 |
ISSN: | 15515044 and 00221554 |
Types: | Journal article |
DOI: | 10.1369/0022155411409411 |
ORCIDs: | Dufva, Martin |
2′-O-methyl RNA Formalin-fixed paraffin-embedded In situ hybridization MicroRNA, locked nucleic acids MicroRNA-138 MicroRNA-205 Urea Yeast RNA
Animals Binding, Competitive Brain Chemistry In Situ Hybridization Mammary Glands, Animal Methylation Mice MicroRNAs Myocardium Nucleic Acid Probes Oligonucleotides RNA Sensitivity and Specificity formalin-fixed paraffin-embedded in situ hybridization locked nucleic acid locked nucleic acids microRNA microRNA-138 microRNA-205 urea yeast RNA